Journal: BMC Bioinformatics

Article Title: Investigating heterogeneous protein annotations toward cross-corpora utilization

doi: 10.1186/1471-2105-10-403

Figure Lengend Snippet: Experimental results of the GENIA subcategory annotations plus AIMed and GENETAG, respectively

Article Snippet: 4 , GENETAG+ DNA , Exact , 69.82 , 36.44 , 47.89.

Techniques:

Journal: Scientific Reports

Article Title: Molecular Combing of Single DNA Molecules on the 10 Megabase Scale

doi: 10.1038/srep19636

Figure Lengend Snippet: ( a ) DNA combing workflow chart. ( b ) DNA combing machine assembled using high precision mechanical modules and adjustable speed. This allows precise movement along the vertical axis with minimal vibration, which helps reduce shearing of DNA molecules. ( c ) Representative image of YOYO-1 stained DNA molecules acquired with epifluorescent microscopy. Four single DNA molecules span the field of view (indicated with arrows).

Article Snippet: Images for DNA molecules were collected in Metamorph (MDS Analytical Technologies) using an epifluorescence microscope (Axioplan 2, Carl Zeiss, Inc) equipped with a Zeiss Plan-FLUAR 63x/1.40 lense (Carl Zeiss, Inc) and CoolSNAP HQ camera (Roper Scientific).

Techniques: Staining, Microscopy

Journal: Scientific Reports

Article Title: Molecular Combing of Single DNA Molecules on the 10 Megabase Scale

doi: 10.1038/srep19636

Figure Lengend Snippet: Fission yeast genomic DNA was labelled in vivo with BrdU using a synchronous cell population. Genomic DNA was prepared for DNA combing from cells at mid S-phase. Single DNA molecules immobilized on glass slide were denatured and BrdU incorporated into newly synthesised DNA was visualised with fluorescently labelled anti-BrdU antibody (green). The whole DNA molecule was counterstained with anti-single stranded DNA antibody (red). ( a ) A representative DNA molecule completely replicated in the presence of BrdU is shown. The quantification of false negative staining, corresponding to the gaps in the BrdU signal expressed as number of pixels is shown below the molecules. Ten DNA molecules corresponding to 5Mb of DNA were analysed. We used a gap size threshold of ≤10 pixels to identify false negative staining in the segmentation of BrdU tracks. ( b ) A representative unlabelled DNA molecule stained with anti-single stranded DNA antibody (red) and anti-BrdU antibody (green) is shown. The quantification of false positive staining, corresponding to green dots on the molecules expressed as number of pixels is shown below the molecule. Ten DNA molecules corresponding to 7 Mb of DNA were analysed. We used a threshold of >3 pixels to identify replication tracks. ( c ) An example of DNA molecules undergoing DNA replication is shown. White arrows point to the middle of replication tracks (shown with a short white line below the DNA molecule) where replication origins are most likely located. The inter origin distance represents the distance between the middle of two adjacent tracks. Biotinylated FISH (fluorescent in situ hybridisation) probes (shown in blue below the molecule) were used to align DNA molecules to their corresponding chromosomal sequences. DNA probes were designed with unique signature left and right arms and a gap between them, allowing the simultaneous detection of different chromosomal loci.

Article Snippet: Images for DNA molecules were collected in Metamorph (MDS Analytical Technologies) using an epifluorescence microscope (Axioplan 2, Carl Zeiss, Inc) equipped with a Zeiss Plan-FLUAR 63x/1.40 lense (Carl Zeiss, Inc) and CoolSNAP HQ camera (Roper Scientific).

Techniques: In Vivo, Negative Staining, Staining, In Situ, Hybridization

Journal: Scientific Reports

Article Title: Molecular Combing of Single DNA Molecules on the 10 Megabase Scale

doi: 10.1038/srep19636

Figure Lengend Snippet: We plotted the cumulative frequencies of inter replication track distances (IRTDs) measured by Patel et al ., 2006 (dark grey squares), the IRTDs measured on full length molecules analysed in our study (black triangles), and the same molecules analysed in our study but “cut” in silico to shorter 500 Kb fragments (light grey circles). The “cut” fragments show a similar distribution of IRTDs as the molecules analysed by Patel et al ., 2006, indicating that the failure to detect clusters of closely spaced origins previously was due to the relatively short size of single DNA molecules analysed.

Article Snippet: Images for DNA molecules were collected in Metamorph (MDS Analytical Technologies) using an epifluorescence microscope (Axioplan 2, Carl Zeiss, Inc) equipped with a Zeiss Plan-FLUAR 63x/1.40 lense (Carl Zeiss, Inc) and CoolSNAP HQ camera (Roper Scientific).

Techniques: In Silico

Journal: Scientific Reports

Article Title: Molecular Combing of Single DNA Molecules on the 10 Megabase Scale

doi: 10.1038/srep19636

Figure Lengend Snippet: ( a ) U2OS cells were synchronised and genomic DNA labelled in vivo by adding BrdU to the media before the onset of S-phase. DNA was prepared for combing from cells at mid-S-phase. BrdU incorporated into newly synthesised DNA was detected with fluorescently labelled anti-BrdU antibody (green) and the entire DNA molecule was counterstained with anti-single stranded DNA antibody (red). To visualise individual replication tracks, the molecule was “cut” in silico into 44 consecutive fragments of 270 kb each, and a composite picture was constructed. A representation of the entire 12 Mb single DNA molecule is shown below, with green and black bars corresponding to replication tracks and to un-replicated segments of the DNA molecule. Black and green bars are drawn to scale. ( b ) Semi-log plot of the cumulative frequencies of IODs for molecules replicated to 50%. The values on the y-axis correspond to the fraction of IODs that are larger in size than the corresponding IOD on the x-axis. The distribution of the experimentally measured values deviates significantly from a purely stochastic distribution, which is expected to fit a straight line (p < 0.001 Lilliefors statistical test). The Lilliefors statistical test is a non-parametric test for exponential distribution, which allows rejection of the hypothesis that the rate of origin firing is homogeneous along the sequence during the early stage of replication, without using any prior estimates from the data, such as the average firing rate. Instead, the cumulative frequencies of IODs lay on a plot with a complex shape that can be decomposed into two straight lines that cross at approximately 40 kb. The straight line with the steeper slope corresponds to IODs for origins firing stochastically at a higher rate within the clusters, whereas the straight line with the shallower slope corresponds to IODs firing stochastically at a lower rate within the regions between clusters (n = 577) ( c ) Histogram of cluster size for molecules replicated from 10% to 50% (45 Mb analysed).

Article Snippet: Images for DNA molecules were collected in Metamorph (MDS Analytical Technologies) using an epifluorescence microscope (Axioplan 2, Carl Zeiss, Inc) equipped with a Zeiss Plan-FLUAR 63x/1.40 lense (Carl Zeiss, Inc) and CoolSNAP HQ camera (Roper Scientific).

Techniques: In Vivo, In Silico, Construct, Sequencing